Biophysical studies of mutated K562 DNA (erythroleukemic cells) binding to adriamycin and daunomycin reveal that mutations induce structural changes influencing binding behavior

Abstract

K562 cells are erythroleukemic cells derived from a chronic myeloid leukemia patient in blast crisis. Comparison of the genome from K562 cells and normal human genome has been very useful strategy, in uncovering eight genes, implicated in acute myeloid leukemia (AML). These genes carry mutations in K562 genome and the role of these mutations in the progression and treatment of AML is still not known. Consequences of these mutations on drug DNA binding are also not known exactly. In the present study, mutation induced structural changes in K562 genome, compared to normal genome, are identified by Fourier transform infra red (FTIR) and circular dichroism (CD) spectroscopy. These structural changes in native K562 DNA favor stronger binding with binding constants 2.0 × 10⁸ and 1.9 × 10⁹ M⁻¹ with antileukemic drugs adriamycin and daunomycin (DNM), respectively, compared to normal DNA. On binding, these drugs disrupt the native B form structure of normal DNA to a greater extent, compared to A-like structure of K562 DNA. Fluorescence and absorption studies reveal higher intercalation as well as mixed groove binding of these drugs with K562 DNA compared to normal DNA. Among the drugs, DNM has higher affinity for K562 DNA.

Mutation Induced Conformational Changes in Genomic DNA from Cancerous K562 Cells Influence Drug-DNA Binding Modes

“…2a–b). Highest binding affinities are associated with M-DNA-DNM interactions, consistent with our earlier findings [6].…”

“…The multiple genetic alterations (mutations) disturb the DNA structure in cancer cells and can also be considered as therapeutic target instead of biochemical manifestations. Such alterations in DNA structure were identified in myeloid leukemic cells (K562), where a partial conformational change from B to A form was reported by us [6].…”

“…These structural changes are evidenced by appearance of new peaks in Raman spectra in N-DNA drug complexes, which are representative of A form DNA (Table 2). M-DNA in native form itself has signatures of A form DNA which has also been consolidated earlier by FTIR and CD [6]. Existing literature on drug DNA interaction theoretical predictions have been made of existence of other forms of associations before final intercalation [44].…”

“…Isolated DNA was further purified by phenol chloroform method and lyophilized [6], [15]. Lyophilized DNA was dissolved in 50 mM phosphate buffer (pH 6.5) when required and the purity was ascertained by the ratio of A 260 /A 280 ; samples with the ratio between of 1.8–2.0 were considered pure.…”

“…These mutations influence the conformation of the DNA and make ADR and DNM binding more effective compared to normal cells. The changes in structure as diagnosed by circular dichroism (CD) and Fourier transform infrared (FTIR) spectroscopy are attributed to these mutations [6], [15]. Such conformational changes and their influence on binding affinities can be thermodynamically characterized.…”